摘要
目的:探讨脓毒症状态下中性粒细胞对T淋巴细胞功能的影响及相关机制。方法:采用脂多糖(lipopolysaccharide,LPS)刺激中性粒细胞,模拟脓毒症状态下的体外模型。分离人外周血中性粒细胞并分离出淋巴细胞,分为对照组、LPS组、中性粒细胞组和LPS+中性粒细胞组4个组。采用流式细胞术检测淋巴细胞凋亡、增殖、活性和炎症细胞因子浓度,以及中性粒细胞表面程序性死亡配体1(PD-L1)的表达;流式细胞术检测使用不同浓度PDL1抗体后的淋巴细胞活性。结果:与对照组相比,中性粒细胞组和LPS+中性粒细胞组淋巴活性明显降低,凋亡明显增加,细胞增殖明显降低,CD69和CD71表达百分比、平均荧光强度、γ干扰素和IL-2的释放显著降低(P均<0. 01);与LPS组相比,LPS+中性粒细胞组能显著抑制淋巴细胞增殖、活化和细胞因子的释放(P均<0. 05);与对照组相比,LPS组PD-L1表达明显增加(P <0. 05);与对照组相比,中、高浓度组淋巴细胞存活百分比明显增加(P均<0. 05),且呈浓度依赖性。结论:脓毒症状态下中性粒细胞通过PD-L1/PD-1通路途径抑制T淋巴细胞的功能。
Objective: To investigate the effect of neutrophils on T lymphocytes during sepsis and its potential mechanism. Methods: Sepsis model in vitro was established by co-culturing T lymphocytes with lipopolysaccharide( LPS). Neutrophils were isolated by density gradient centrifugation and T lymphocytes were isolated by anti-CD3 magnetic beads from human peripheral blood. T lymphocytes were divided into4 groups,including control group,LPS group,neutrophil group and LPS + neutrophil group. The apoptosis,proliferation,activation and secretion of T lymphocytes were assessed by flow cytometry. Phenotype programmed death-ligand 1( PD-L1) on neutrophil was detected in control group and LPS group. AntiPD-L1 antibody was added into mixed culture of neutrophils with T cells and the apoptosis of T cells were assessed. Results: Compared with the control group,neutrophil group and LPS + neutrophil group showed significantly lower immunoactivity of T lymphocytes,higher proportion of apoptosis,lower expression levels of CD69 and CD71,as well as secretion of interferon-γ( IFN-γ) and IL-2( all P < 0. 05).Compared with LPS group,the proliferation of T cells,secretion of IFN-γ and IL-2 were decreased in LPS + neutrophilgroup( all P < 0. 05). The ratio of PD-L1 in LPS group was significantly higher than that of the control group( P < 0. 05). The proportion of viable lymphocytes increased with higher concentrations of PD-L1 monoclonal antibody( all P < 0. 05). Conclusion: Neutrophils suppressed the immuno-activity of T lymphocytes through PD-L1/PD-1 pathway during sepsis.
作者
于垚
孙辉
柳益书
戚欣欣
刘璐
孙炳伟
YU Yao;SUN Hui;LIU Yi-shu;QI Xin-xin;LIU Lu;SUN Bing-wei(School of Medicine,Jiangsu University,Zhenjiang Jiangsu 212013;Department of Burns and Plastic Surgery,Suzhou Hospital Affiliated to Nanjing Medical University,Suzhou Jiangsu 215008,China)
出处
《江苏大学学报（医学版）》
CAS
2018年第6期467-472,共6页
Journal of Jiangsu University：Medicine Edition
基金
国家自然科学基金资助项目(81772135
81471903)
作者简介
于垚(1993—),女,硕士研究生;孙炳伟(通讯作者),主任医师,教授,博士生导师,E-mail:sunbinwe@hotmail.com
